A. Structure-Function Analysis of cAMP-independent Forms of the cAMP Receptor Protein (CRP). The cAMP receptor protein (CRP) ordinarily requires cAMP for function in operon expression. Mutants of CRP, termed CRP* proteins, have the unique property of functioning in the absence of added cAMP. Sequence analysis of cloned CRP* DNA has allowed the primary structure of such proteins to be determined and compared with that of the wild-type protein. The gene for one of the cAMP-independent proteins, termed NCR91, has been cloned into a high expression vector; using this construct, it has been possible to grow E. coli under conditions where substantial amounts of CRP91 are formed. The protein has been crystallized and its structure analyzed by X-ray diffraction. This data has permitted a comparison of the crystal structure of the mutant protein with that of the wild-type protein. B. Regulation of Sugar Transport in Gram-Positive Bacteria. Regulation of sugar metabolism in gram-positive bacteria is uniquely different from that in gram-negative bacteria, because gram-positive bacteria don't make cAMP. In gram-negative bacteria, the adenylate cyclase system is regulated by the phosphoenolpyruvate:sugar phosphotransferase system (PTS). Some gram-positive bacteria transport sugars by the PTS; this sugar transport system also plays a role in the process of inducer expulsion whereby the addition of glucose to cells loaded with a nonmetabolizable sugar triggers the elimination of that sugar. Some strains of the gram-positive lactobacilli do not have an active PTS but nevertheless show the process of inducer expulsion. The characteristics of the inducer expulsion process in Lactobacillus brevis was studied and it was demonstrated that, although there is not a functional PTS in this organism, one of the components of the sugar transport system is present. It is suggested that this component may play a role in regulating sugar transport in Lactobacillus brevis.